PROJECT SUMMARY: Approximately 70% of American adults are considered overweight or obese putting them at increased risk for metabolic complications such as non-alcoholic fatty liver disease (NAFLD). NAFLD encompasses a spectrum of steatogenic (i.e. increased liver triglycerides) liver disorders that lead to an increased risk for type II diabetes, insulin resistance and obesity-related mortality. Sexual dimorphism is clearly established, with the incidence of NAFLD being significantly higher in males than premenopausal females. The mechanisms that protect females from fatty liver remain unclear. While the majority of investigations have taken a liver-centric perspective, our recent findings indicate that endoplasmic reticulum stress in the subfornical organ (SFO), a forebrain circumventricular region that senses circulating factors, mediates hepatic steatosis during obesity in male mice. Importantly, the SFO has dense excitatory projections to the paraventricular nucleus of the hypothalamus (PVN), an integrative nucleus that plays a critical role in autonomic and endocrine control and has direct spinal projections to the liver. Our exciting data indicate that activation of SFO neurons that project to the PVN (hereafter referred to as SFOPVN neurons) results in elevations in hepatic sympathetic outflow that are accompanied by the development of NAFLD. Furthermore, inhibition of this forebrain-hypothalamic circuit or removal of the hepatic sympathetic nerves is sufficient to rescue obesity-mediated NAFLD. Using a combination of innovative imaging, molecular, neuroanatomical, and integrative physiological approaches, we will build upon these findings and address the central hypothesis that estrogen signaling in SFOPVN projecting neurons protects against NAFLD development in females. Given: 1) Accumulating evidence that estrogen signaling protects against endoplasmic reticulum stress; and 2) Estrogen receptor alpha (ERa) is expressed on SFOPVN projecting neurons, in Aim 1 we will comprehensively evaluate whether estrogen protects against high fat diet-induced endoplasmic reticulum stress in SFOPVN projecting neurons. In line with this, in Aim 2, we will examine whether estrogen signaling in SFOPVN projecting neurons protects against obesity-induced hepatic sympathetic overactivity and NAFLD via endoplasmic reticulum stress mechanisms. Dissecting the links between central nervous system estrogen signaling and endoplasmic reticulum stress in the control of hepatic sympathetic outflow and NAFLD development will advance our understanding of underlying mechanisms that contribute to sex differences in NAFLD. These findings may also help identify new therapeutic targets for treatment and prevention of this condition. Moreover, the combination of elegant intersectional viral targeting strategies, high- resolution microscopy approaches, direct recording of sympathetic outflow to the liver, and comprehensive histological/molecular hepatic analysis provides a novel and integrative approach to gain insight into sexually dimorphic NAFLD development.